Abnormalities of proliferative and differentiative properties in DM1 and DM2 senescence myoblasts -in vitro-

Myotonic dystrophy (DM) is an autosomal dominant multisystemic disorder characterized by a variety of multisystemic features including myotonia, muscular dystrophy, cardiac dysfunctions, cataracts and insulin-resistance. One form of the disorder named myotonic dystrophy type 1 (DM1) is caused by an expanded (CTG)n, in the 3’ untranslated region of the DMPK gene, while a second form of DM (DM2) is caused by the expansion of a tetranucleotidic repeat (CCTG)n in the intron 1 of the ZNF9 gene. The mutant transcripts accumulate in nuclear foci altering the function of the alternative splicing regulators CUGBP1 and MBLN1 proteins, which are necessary for the physiological processing of mRNAs. However, at the skeletal muscle level, this proposed pathogenetic mechanism does not explain the muscle weakness and atrophy observed in DM patients or the muscle histopathological features characteristic of these diseases. It has been observed that DM muscle shares similarities with the ageing muscle, where the progressive muscle weakness and atrophy is accompanied by a slower regenerative capacity possibly due to the failure in satellite cells activation. The aim of this study is to investigate the possible relationship between a premature cell ageing process and a differentiation defect in DM patients and their muscle cell phenotypes. Primary cultures of human myoblasts from DM1 (N=3), DM2 (n=3) and control patients (n=3) have been used to study the replicative in vitro senescence in term of morphology, proliferative and differentiation capability. The expression of several markers of proliferation, differentiation and senescence (PCNA, MyoD, Myogenin, p16…) has been analyzed. BrdU staining, PCNA expression and the mean population doubling values indicate that in vitro senescence of DM1 and DM2 myoblasts results in a reduction in their proliferative capability as compared to control myoblasts. Moreover, during in vitro senescence, a decrease in fusion index and an increase of p16 expression is observable in both DM and control myoblast. This impairment in differentiation potential of senescent myoblasts is more evident in DM muscle cells. The results of this study suggest that alterations in proliferation potential and differentiation capabilities of satellite cells obtained from DM muscle might contribute to some of the clinical and histopathological features observed in DM patients.